In this project, we intend to investigate several questions about Na+ transport in the rabbit urinary bladder. These questions are: (1) What are the characteristics of the Na+ transport system in the basolateral or serosal facing membrane? We will describe the ATP dependence of the basolateral Na+ pump. We will also determine the stoichiometry of the pump and if the stoichiometry is variable. To perform these experiments, we will gain access to the basolateral membrane by permeabilizing the apical membrane with digitonin. Digitonin produces pores in the apical membrane which effectively removes the apical membrane as a permeability barrier to molecules as large as small proteins. (2) Can the dependence of the basolateral Na+ pump on ATP or ADP be an important control factor in regulating transepithelial Na+ transport; i.e., are the normal physiological concentrations of ATP or ADP in a range where the pump is running at less than maximal rate? (3) Are there other intracellular factors which might control the basolateral Na+ pump? In particular, can intracellular H+ ion control the basolateral Na+ transport? (4) Are there other transport mechanisms for moving electrolytes across the basolateral membrane? Specifically, is there Na-C1 co-transport or Na-HCO3 co-transport? (5) Does intracellular Ca++ or H+ control the amiloride-sensitive Na+ conductance of the apical membrane? We will examine this question in a simple model system, the A6 epithelial tissue culture line. The purpose of the above experiments is to define the cellular mechanisms by which Na+ transport is controlled and to develop a model of the transport in detail which has only previously been possible in much simpler tissues.